Neurotransmitters are packaged into synaptic vesicles by protein transporters located in the membrane of these vesicles. Transport activity from these vesicular neurotransmitter transporters was originally characterized biochemically. cDNAs for all classical neurotransmitters, except for the major excitatory neurotransmitter glutamate, have now been identified. The Edwards lab has recently identified a putative vesicular transporter cDNA (vt2) with homology to the vesicular GABA/glycine transporter. From in situ analysis, the vt2 RNA is localized to neurons with particularly dense expression in the hippocampus and the granular cells of the cerebellum. This localization is consistent with a function for the protein in glutamatergic neurons. In addition, localization by immunofluorescence of transfected cell cultures shows an intracellular localization of the protein. These observations are consistent with VT2 being a putative vesicular transporter of glutamate. The goal of this fellowship is to determine the function of the VT2 protein and the physiological role it plays in the animal. Two reagents are being generated in order to analyze this protein: an antibody and a mouse with a targeted disruption in the vt2 gene. The antibody will be used to determine the cellular and subcellular localization of the VT2 protein, whereby directing the type of assay used to subsequently determine its function. And, from recently generated chimeric mice, heterozygous and homozygous mice will be bred and analyzed for any obvious developmental or behavioral phenotypes. These mutant mice will also be used to determine the function of the VT2 protein and the effects that deficiencies have on physiological functions.